Experiments proposed in this application will determine the importance of the Cone-Rod Homeobox (CRX) and its associated regulatory network in photoreceptor gene expression and photoreceptor degenerative diseases. CRX is a novel member of otd/Otx homeodomain transcription factor family that is specifically expressed in photoreceptors. CRX binds to and trans-activates the promoters of many photoreceptor-specific genes. Mutations in CRX are associated with photoreceptor degenerative diseases having a wide age of onset, including cone-rod dystrophy, retinitis pigmentosa and Leber's congenital amaurosis. Our hypothesis is that CRX serves as one-of the key molecules that control the expression of photoreceptor-specific genes and is essential for the normal development and maintenance of the photoreceptors. Two specific aims are proposed to test this hypothesis: Aim I will test how disease-related CRX mutations alter the way that CRX regulates photoreceptor gene expression. Five known CRX homeodomain mutations will be analyzed for their effects on the binding of CRX to DNA and trans-activation of the rhodopsin promoter. Aim 2 will identify and characterize other components of the CRX regulatory pathway, such as proteins that regulate the activity of CRX or co-factors required for CRX function. These proteins are hypothesized to physically interact with CRX and, therefore, will be isolated using protein-protein interaction screens in yeast cells. The resulting positive clones will be characterized by sequence homology to known proteins, expression patterns, and physical and functional interactions with CRX. We will also determine the chromosomal localization of the corresponding human and murine genes. These studies will provide information about the program controlling photoreceptor-specific gene expression and the development and maintenance of photoreceptor function. They are expected to provide information about the molecular basis of photoreceptor degenerative diseases and aid in the selection of targets for therapeutic approaches to these diseases.